摘要
目的 探讨羟基脲(hydroxyurea,HU)对密闭缺氧小鼠的保护作用及机制。方法 将60只6~8周龄、体质量18~22g的雄性C57BL/6J小鼠随机分成常氧对照组(NC组,n=10)、常氧HU组(NHU组,n=10)、缺氧对照组(HC组,n=20)、缺氧HU组(HHU组,n=20)。HU组每天腹腔注射HU100mg/kg;对照组每天注射等量生理盐水。4周后取NHU组和NC组小鼠的骨髓进行转录组测序;再将HHU组和HC组置于密封瓶,进行密闭缺氧,记录其生存时间;在缺氧15min时,取脑、肾组织进行免疫组化,同时取未缺氧的NHU组和NC组脑、肾组织进行对照;并对骨髓进行RT-qPCR验证,外周血进行Westernblot验证。结果 HHU组小鼠密闭缺氧的生存时间显著延长(P<0.05);脑和肾脏缺氧程度显著降低(P<0.05)。外周血氧亲和力显著提高,且P50与小鼠生存时间存在显著的负相关关系(Pearson=-0.738,P<0.01)。转录组共检测到65389个基因,其中NHU组上调基因50个,下调基因272个(|log2FC|≥1,Q<0.05),对差异显著的基因进行功能富集,结果发现在分子组成上,血红蛋白复合体居于首位;分子功能上,主要富集到影响血红蛋白各亚基结合的分子;生物学功能上主要富集到红细胞发育相关的生物学过程。差异分子中,Erfe、Hbb-y、Hbb-bh1、Hba-x>1.5倍以上,而下调基因有红系发育相关基因Klf1、Sox6、Gata1等;经验证,NHU组Hbb-y、Hbb-bh1、Hba-x的mRNA显著升高,外周血HbE(ε-globin)蛋白表达增加,差异具有统计学意义(P<0.05)。结论 HU对密闭缺氧小鼠具有明显的保护作用,可能与HU诱导高氧亲和力的胚胎样血红蛋白表达,从而促进携氧的作用有关。
Objective To investigate the ameliorative effect and mechanism of hydroxyurea(HU) on hypoxia mice in a closed environment.Methods Sixty male C57BL/6J mice(6~8 weeks old,weighing 18~22 g) were randomly divided into normoxia control group(NC,n=10),normoxia HU group(NHU,n=10),hypoxia control group(HC,n=20),and hypoxia HU group(HHU,n=20).The HU mice were injected intraperitoneally with 100 mg/kg HU daily for 4 weeks,and the control mice were injected with same volume saline for 4 weeks.Then bone marrow was extracted for transcriptome sequence in the NC and NHU groups.The mice in the HC and HHU groups were settled in a sealed bottle to observe their survival,and the brains and kidneys were isolated for immunohistochemistry when these mice were in hypoxia for 15 min,and those from the NHU and NC groups served as control.RT-qPCR and Western blotting were adopted to verify the mRNA and protein expression of bone marrow and peripheral blood molecules,respectively.Results The survival time was significantly longer in the HHU group(P<0.05),and the hypoxia of the brains and kidneys were ameliorated(P<0.05).The oxygen affinity of peripheral blood was obviously raised,and partial pressure of oxygen 50(P50) was negatively correlated with survival time(Pearson=-0.738,P<0.01).A total of 65 389 genes were found in transcription sequencing,including 50 up-regulated and 272 down-regulated genes in the mice from the NHU group (|log2FC|≥1,Q<0.05).Gene ontology(GO) enrichment analysis indicated that hemoglobin subunits(GO cellular component),hemoglobin complex(GO cellular function) and erythrocyte development(GO biological process) were significant enriched.Among the differentially expressed genes,Erfe,Hbb-y,Hbb-bh1 and Hba-x were up to 1.5 fold,and the down-regulated genes were erythropoiesis-related genes,Klf1,Sox6,Gata1 etc.Our results indicated that the mRNA levels of Hbb-y,Hbb-bh1 and Hba-x were notably enhanced in the bone marrow of the NHU group,and the protein level of HbE(ε-globin) protein was remarkably increased in the pe
作者
周晓英
矫力
崔宇
李晓栩
阳一栋
杨诚忠
王辰元
黄缄
ZHOU Xiaoying;JIAO Li;CUI Yu;LI Xiaoxu;YANG Yidong;YANG Chengzhong;WANG Chenyuan;HUANG Jian(Department of Plateau Physiology and Pathology,Faculty of High Altitude Military Medicine,Army Medical University(Third Military Medical University),Chongqing,400038,China)
出处
《陆军军医大学学报》
CAS
CSCD
北大核心
2023年第2期93-101,共9页
Journal of Army Medical University
基金
军队后勤科研项目(BWS17J031)
军队医学科技青年培育计划(20QNPY002)。
